Project Summary Myelination of the central nervous system (CNS) by oligodendrocytes (OLs) is essential for its development and function. Myelin sheaths develop in the CNS as OLs undergo terminal differentiation, implicating OL differentiation as a crucial event for CNS myelination. OL differentiation also represents a key rate-limiting step of remyelination in pathological conditions. Transcriptional regulation plays a pivotal role in the differentiation of OLs, which is a team effort of trans-acting factors and cis-regulatory DNA elements such as enhancers. Much work has been devoted to trans-acting factors, identifying important transcription factors for OL development. In contrast, relatively little is known about enhancers that orchestrate the differentiation of OLs. Further, the role of OL enhancer variants in the etiology of CNS disorders remains poorly understood. Building upon our innovative preliminary study, this grant intends to address these critical issues. Sox10 is a high-mobility group transcription factor essential for OL differentiation and CNS myelination. Yet, the identity of OL enhancers that regulate Sox10 expression remains elusive. Aim I intends to reveal an intricate network of OL enhancers that governs Sox10 expression for timely OL differentiation. We found that 75 single nucleotide polymorphisms (SNPs) associated with CNS traits are mapped to putative OL enhancers, prompting us to hypothesize that CNS trait SNPs may exert their effects by impacting OL enhancers and the property of OL lineage cells. As a proof of principle, Aim II will examine whether this is true for 3 CNS trait SNPs. In sum, this study will elucidate the understudied, yet crucial role of OL enhancers in CNS development and disease.